Nature's approach toward ring formation and structural diversity in ergot alkaloid biosynthesis

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Chemistry, 2011.Vita. Cataloged from PDF version of thesis.Includes bibliographical references.Ergot alkaloids are fungal-derived secondary metabolites well known for a diverse array of pharmacological effects both beneficial and detrimental to human health. Historically, the ergot alkaloids have been known to cause ergotism in populations that consume grain contaminated by ergot alkaloid producing fungus. However, naturally isolated and semi-synthetic derivatives of certain ergot alkaloids have also been used to treat migraines, Parkinsonism, and tumor growth. Different fungal species such as Aspergillusfumigatus, Claviceps purpurea, and Neotyphodium lolii produce ergot alkaloids that have distinct structural features yet share a common tetracyclic ergoline ring scaffold. Mechanistic details regarding the formation of the common ergoline ring are not well understood, though the genes encoding the enzymes that carry out these cyclizations are believed to be well conserved across divergent fungal species. Here we describe in vivo gene disruption experiments in Aspergillusfumigatus that allowed us to identify candidate enzymes that were directly involved with the intramolecular cyclization of the ergoline ring. Additionally, we discuss the cloning and heterologous expression of these genes to further characterize their catalytic function. Old Yellow Enzyme homologues from the ergot gene clusters of Aspergillus fumigatus and Neotyphodium loii were shown to catalyze the formation of the D ring of the ergoline skeleton. These enzymes catalyzed either reductase or isomerase type reactions to yield distinct pathway intermediates. Mutational analysis was used to engineer an Old Yellow Enzyme that displayed both reductase and isomerase activities, thereby elucidating the mechanistic basis behind this switch in enzymatic activity. These findings present a mechanistic rationale behind nature's biosynthetic strategy toward ring cyclization and the introduction of structural diversity into the ergot alkaloid class of natural products.by Johnathan Zandrew Cheng.Ph.D

18F-FDG-PET/CT in Radiation Therapy-Induced Cerebellar Inflammation

ABSTRACT Background 18F-fluorodeoxyglucose-positron emission tomography/computed tomography (18FDG- PET/CT) is used in the clinical diagnosis and management of oncologic and inflammatory pathologies. It may also have utility in detecting tissue damage induced by radiotherapy (RT) used to treat various types of cancer. The aim of the present study was to use 18FDG-PET/CT to evaluate the effect of RT on the uptake of 18FDG by the cerebellum. Methods Thirty patients with head and neck cancer (HNC) were included in this retrospective study. The patients were treated with photon, proton, or combined photon/proton RT, in addition to chemotherapy. All patients received 18FDG- PET/CT imaging pre-treatment and 3 months post-treatment. The global mean standardized uptake value (SUVmean) of the cerebellum was determined for every patient by global assessment of 18FDG activity using OsiriX MD software. A two-tailed paired t-test was used to compare global SUVmean pre- and post-RT. Results The pre-treatment and post-treatment global SUVmean for the photon group were 5.26 and 5.51 (p: 0.42), respectively. As for the proton only group, the pre- and post-treatment global SUVmeans were 7.06 and 6.05, respectively. In the combined RT group, the pre- and post-treatment global SUVmeans were 6.14 and 6.19 respectively (p: 0.92). The differences between the pre- and post-treatment values failed to reach statistical significance for any of the treatment groups but it should be noted that there was a trend of increased 18FDG uptake in the cerebellum following photon therapy. This trend was not clear in the combined group. As for the proton group, p-value was not calculated as only two patients were included. Conclusion Although not statistically significant, the results showed an incremental increase in global SUVmean following treatment with photon RT likely reflecting the presence of mild radiation-induced inflammation in the cerebellum

The effect of novel synthetic methods and parameters control on morphology of nano-alumina particles

Alumina is an inorganic material, which is widely used in ceramics, catalysts, catalyst supports, ion exchange and other fields. The micromorphology of alumina determines its application in high tech and value-added industry and its development prospects. This paper gives an overview of the liquid phase synthetic method of alumina preparation, combined with the mechanism of its action. The present work focuses on the effects of various factors such as concentration, temperature, pH, additives, reaction system and methods of calcination on the morphology of alumina during its preparation

Dynamically Controlling Image Integration Onboard the Star-Planet Activity Research CubeSat (SPARCS)

The Star-Planet Activity Research CubeSat (SPARCS) is a 6U CubeSat astronomical observatory underdevelopment and will be entirely dedicated to the photometric monitoring of the flaring activity of M dwarfs at near-UV (258 nm – 308 nm) and far-UV (153 nm–171 nm) wavelengths. The SPARCS science pay load is composed of a 9-cm telescope that projects a 40’ field-of-view onto two UV-optimized delta-doped charge-coupled devices (CCDs), which are controlled by a dedicated payload processor board. Given that M dwarf flares in the UV are expected to be capable of reaching amplitudes ∼14,000 times above their quiescent flux, with durations that can be as short as a couple of minutes, the SPARCS payload processor is designed to be able to dynamically adjust the imaging system’s integration times and gains on the fly to reduce CCD pixel saturation issues when flaring events are detected. The SPARCS payload processor is a BeagleBone Black (BBB) with a protective Pumpkin Motherboard Module 2, and runs a custom fully Python-based software to perform active detector thermal control, manage science observations, and apply near-real time image processing to autonomously adjust the exposure times and gains of the detectors upon flare detection. Here we present the approach adopted for that automated dynamic exposure control, as well as its pre-flight tests and performance using simulated M dwarf light curves and full-frame images in the two SPARCS passbands

Prediction of protein assemblies, the next frontier: The CASP14-CAPRI experiment

We present the results for CAPRI Round 50, the fourth joint CASP-CAPRI protein assembly prediction challenge. The Round comprised a total of twelve targets, including six dimers, three trimers, and three higher-order oligomers. Four of these were easy targets, for which good structural templates were available either for the full assembly, or for the main interfaces (of the higher-order oligomers). Eight were difficult targets for which only distantly related templates were found for the individual subunits. Twenty-five CAPRI groups including eight automatic servers submitted ~1250 models per target. Twenty groups including six servers participated in the CAPRI scoring challenge submitted ~190 models per target. The accuracy of the predicted models was evaluated using the classical CAPRI criteria. The prediction performance was measured by a weighted scoring scheme that takes into account the number of models of acceptable quality or higher submitted by each group as part of their five top-ranking models. Compared to the previous CASP-CAPRI challenge, top performing groups submitted such models for a larger fraction (70–75%) of the targets in this Round, but fewer of these models were of high accuracy. Scorer groups achieved stronger performance with more groups submitting correct models for 70–80% of the targets or achieving high accuracy predictions. Servers performed less well in general, except for the MDOCKPP and LZERD servers, who performed on par with human groups. In addition to these results, major advances in methodology are discussed, providing an informative overview of where the prediction of protein assemblies currently stands.Cancer Research UK, Grant/Award Number: FC001003; Changzhou Science and Technology Bureau, Grant/Award Number: CE20200503; Department of Energy and Climate Change, Grant/Award Numbers: DE-AR001213, DE-SC0020400, DE-SC0021303; H2020 European Institute of Innovation and Technology, Grant/Award Numbers: 675728, 777536, 823830; Institut national de recherche en informatique et en automatique (INRIA), Grant/Award Number: Cordi-S; Lietuvos Mokslo Taryba, Grant/Award Numbers: S-MIP-17-60, S-MIP-21-35; Medical Research Council, Grant/Award Number: FC001003; Japan Society for the Promotion of Science KAKENHI, Grant/Award Number: JP19J00950; Ministerio de Ciencia e Innovación, Grant/Award Number: PID2019-110167RB-I00; Narodowe Centrum Nauki, Grant/Award Numbers: UMO-2017/25/B/ST4/01026, UMO-2017/26/M/ST4/00044, UMO-2017/27/B/ST4/00926; National Institute of General Medical Sciences, Grant/Award Numbers: R21GM127952, R35GM118078, RM1135136, T32GM132024; National Institutes of Health, Grant/Award Numbers: R01GM074255, R01GM078221, R01GM093123, R01GM109980, R01GM133840, R01GN123055, R01HL142301, R35GM124952, R35GM136409; National Natural Science Foundation of China, Grant/Award Number: 81603152; National Science Foundation, Grant/Award Numbers: AF1645512, CCF1943008, CMMI1825941, DBI1759277, DBI1759934, DBI1917263, DBI20036350, IIS1763246, MCB1925643; NWO, Grant/Award Number: TOP-PUNT 718.015.001; Wellcome Trust, Grant/Award Number: FC00100

Evaluating the Effects of SARS-CoV-2 Spike Mutation D614G on Transmissibility and Pathogenicity.

Global dispersal and increasing frequency of the SARS-CoV-2 spike protein variant D614G are suggestive of a selective advantage but may also be due to a random founder effect. We investigate the hypothesis for positive selection of spike D614G in the United Kingdom using more than 25,000 whole genome SARS-CoV-2 sequences. Despite the availability of a large dataset, well represented by both spike 614 variants, not all approaches showed a conclusive signal of positive selection. Population genetic analysis indicates that 614G increases in frequency relative to 614D in a manner consistent with a selective advantage. We do not find any indication that patients infected with the spike 614G variant have higher COVID-19 mortality or clinical severity, but 614G is associated with higher viral load and younger age of patients. Significant differences in growth and size of 614G phylogenetic clusters indicate a need for continued study of this variant

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Hospital admission and emergency care attendance risk for SARS-CoV-2 delta (B.1.617.2) compared with alpha (B.1.1.7) variants of concern: a cohort study

Background: The SARS-CoV-2 delta (B.1.617.2) variant was first detected in England in March, 2021. It has since rapidly become the predominant lineage, owing to high transmissibility. It is suspected that the delta variant is associated with more severe disease than the previously dominant alpha (B.1.1.7) variant. We aimed to characterise the severity of the delta variant compared with the alpha variant by determining the relative risk of hospital attendance outcomes. Methods: This cohort study was done among all patients with COVID-19 in England between March 29 and May 23, 2021, who were identified as being infected with either the alpha or delta SARS-CoV-2 variant through whole-genome sequencing. Individual-level data on these patients were linked to routine health-care datasets on vaccination, emergency care attendance, hospital admission, and mortality (data from Public Health England's Second Generation Surveillance System and COVID-19-associated deaths dataset; the National Immunisation Management System; and NHS Digital Secondary Uses Services and Emergency Care Data Set). The risk for hospital admission and emergency care attendance were compared between patients with sequencing-confirmed delta and alpha variants for the whole cohort and by vaccination status subgroups. Stratified Cox regression was used to adjust for age, sex, ethnicity, deprivation, recent international travel, area of residence, calendar week, and vaccination status. Findings: Individual-level data on 43 338 COVID-19-positive patients (8682 with the delta variant, 34 656 with the alpha variant; median age 31 years [IQR 17–43]) were included in our analysis. 196 (2·3%) patients with the delta variant versus 764 (2·2%) patients with the alpha variant were admitted to hospital within 14 days after the specimen was taken (adjusted hazard ratio [HR] 2·26 [95% CI 1·32–3·89]). 498 (5·7%) patients with the delta variant versus 1448 (4·2%) patients with the alpha variant were admitted to hospital or attended emergency care within 14 days (adjusted HR 1·45 [1·08–1·95]). Most patients were unvaccinated (32 078 [74·0%] across both groups). The HRs for vaccinated patients with the delta variant versus the alpha variant (adjusted HR for hospital admission 1·94 [95% CI 0·47–8·05] and for hospital admission or emergency care attendance 1·58 [0·69–3·61]) were similar to the HRs for unvaccinated patients (2·32 [1·29–4·16] and 1·43 [1·04–1·97]; p=0·82 for both) but the precision for the vaccinated subgroup was low. Interpretation: This large national study found a higher hospital admission or emergency care attendance risk for patients with COVID-19 infected with the delta variant compared with the alpha variant. Results suggest that outbreaks of the delta variant in unvaccinated populations might lead to a greater burden on health-care services than the alpha variant. Funding: Medical Research Council; UK Research and Innovation; Department of Health and Social Care; and National Institute for Health Research

Evaluating the Effects of SARS-CoV-2 Spike Mutation D614G on Transmissibility and Pathogenicity

Global dispersal and increasing frequency of the SARS-CoV-2 spike protein variant D614G are suggestive of a selective advantage but may also be due to a random founder effect. We investigate the hypothesis for positive selection of spike D614G in the United Kingdom using more than 25,000 whole genome SARS-CoV-2 sequences. Despite the availability of a large dataset, well represented by both spike 614 variants, not all approaches showed a conclusive signal of positive selection. Population genetic analysis indicates that 614G increases in frequency relative to 614D in a manner consistent with a selective advantage. We do not find any indication that patients infected with the spike 614G variant have higher COVID-19 mortality or clinical severity, but 614G is associated with higher viral load and younger age of patients. Significant differences in growth and size of 614G phylogenetic clusters indicate a need for continued study of this variant

Changes in symptomatology, reinfection, and transmissibility associated with the SARS-CoV-2 variant B.1.1.7: an ecological study

Background The SARS-CoV-2 variant B.1.1.7 was first identified in December, 2020, in England. We aimed to investigate whether increases in the proportion of infections with this variant are associated with differences in symptoms or disease course, reinfection rates, or transmissibility. Methods We did an ecological study to examine the association between the regional proportion of infections with the SARS-CoV-2 B.1.1.7 variant and reported symptoms, disease course, rates of reinfection, and transmissibility. Data on types and duration of symptoms were obtained from longitudinal reports from users of the COVID Symptom Study app who reported a positive test for COVID-19 between Sept 28 and Dec 27, 2020 (during which the prevalence of B.1.1.7 increased most notably in parts of the UK). From this dataset, we also estimated the frequency of possible reinfection, defined as the presence of two reported positive tests separated by more than 90 days with a period of reporting no symptoms for more than 7 days before the second positive test. The proportion of SARS-CoV-2 infections with the B.1.1.7 variant across the UK was estimated with use of genomic data from the COVID-19 Genomics UK Consortium and data from Public Health England on spike-gene target failure (a non-specific indicator of the B.1.1.7 variant) in community cases in England. We used linear regression to examine the association between reported symptoms and proportion of B.1.1.7. We assessed the Spearman correlation between the proportion of B.1.1.7 cases and number of reinfections over time, and between the number of positive tests and reinfections. We estimated incidence for B.1.1.7 and previous variants, and compared the effective reproduction number, Rt, for the two incidence estimates. Findings From Sept 28 to Dec 27, 2020, positive COVID-19 tests were reported by 36 920 COVID Symptom Study app users whose region was known and who reported as healthy on app sign-up. We found no changes in reported symptoms or disease duration associated with B.1.1.7. For the same period, possible reinfections were identified in 249 (0·7% [95% CI 0·6–0·8]) of 36 509 app users who reported a positive swab test before Oct 1, 2020, but there was no evidence that the frequency of reinfections was higher for the B.1.1.7 variant than for pre-existing variants. Reinfection occurrences were more positively correlated with the overall regional rise in cases (Spearman correlation 0·56–0·69 for South East, London, and East of England) than with the regional increase in the proportion of infections with the B.1.1.7 variant (Spearman correlation 0·38–0·56 in the same regions), suggesting B.1.1.7 does not substantially alter the risk of reinfection. We found a multiplicative increase in the Rt of B.1.1.7 by a factor of 1·35 (95% CI 1·02–1·69) relative to pre-existing variants. However, Rt fell below 1 during regional and national lockdowns, even in regions with high proportions of infections with the B.1.1.7 variant. Interpretation The lack of change in symptoms identified in this study indicates that existing testing and surveillance infrastructure do not need to change specifically for the B.1.1.7 variant. In addition, given that there was no apparent increase in the reinfection rate, vaccines are likely to remain effective against the B.1.1.7 variant. Funding Zoe Global, Department of Health (UK), Wellcome Trust, Engineering and Physical Sciences Research Council (UK), National Institute for Health Research (UK), Medical Research Council (UK), Alzheimer's Society